Flagellin, a TLR5 Agonist, Reduces Graft-versus-Host Disease in Allogeneic Hematopoietic Stem Cell Transplantation Recipients While Enhancing Antiviral Immunity

Abstract

Graft-versus-host disease (GvHD) is a major cause of morbidity and mortality in patients treated with allogeneic hematopoietic stem cell transplantation (HSCT). Post-transplant immunosuppressive drugs incompletely control GvHD and increase susceptibility to opportunistic infections. In this study we used flagellin, a TLR5 agonist protein (~50 kDa) extracted from bacterial flagella, as a novel experimental treatment strategy to reduce both acute and chronic GvHD in allogeneic HSCT recipient. Based upon the radio-protective effects of flagellin, we hypothesized that flagellin could ameliorate GvHD in lethally irradiated murine models of allogeneic HSCT. Two doses of highly purified flagellin (administered 3 hrs. before irradiation and 24 hrs. after HSCT) reduced GvHD and led to better survival in both H-2b → CB6F1 and H-2k → B6 allogeneic HSCT models while preserving over 99% donor T cells chimerism. Flagellin treatment preserved long-term post-transplant immune reconstitution characterized by more donor thymic-derived CD4+CD25+foxp3+ regulatory T cells (Tregs) and significantly enhanced anti-viral immunity following murine cytomegalovirus (mCMV) infection model. The proliferation index and activation status of donor spleen-derived T cells, and serum concentration of pro-inflammatory cytokines in flagellin-treated recipients were reduced significantly within 4 days post-transplant compared with the PBS-treated control recipients. Allogeneic transplantation of radiation chimeras previously engrafted with TLR5 knockout hematopoietic cells showed that interactions between flagellin and TLR5 expressed on both donor hematopoietic and host non-hematopoietic cells were required to reduce GvHD. Thus, the peri-transplant administration of flagellin is a novel therapeutic approach control GvHD while preserving post-transplant donor immunity.

A TLR5 Agonist Enhances CD8+ T Cell-Mediated Graft-versus-Tumor Effect without Exacerbating Graft-versus-Host Disease

“…Therefore, it is a significant finding that CBLB502 treatment enhances GVT effect but does not cause more severe GVHD. However, our results are not entirely consistent with a recent report, which used tumor-free hosts to show that peritransplant administration of flagellin reduced GVHD in an allo-BMT model whereas enhancing donor-derived antiviral immunity (44). The discrepancy may be because of several factors, besides the difference between flagellin and CBLB502.…”

Recombinant TLR5 Agonist CBLB502 Promotes NK Cell-Mediated Anti-CMV Immunity in Mice

“…Paradoxically, flagellin treatment also protected allo-BMT recipients from lethal mCMV infection [8]. Like native flagellin, rflagellin also protected allo-BMT recipients from GvHD in a similar fashion, with the optimal i.p dose between 25 µg to 50 µg/mouse (our unpublished data).…”

“…Thus, the optimal schedule of rflagellin administration, 2 days before viral infection, might preclude its clinical application as treatment or prophylaxis for mCMV infection. However, administration of rflagellin significantly reduced production of inflammatory cytokines on day 0 of mCMV infection (Figure 8G and H), and peritransplant administration of native flagellin reduced GvHD and also reduced inflammatory cytokines in allo-BMT recipients [8]. Since inflammatory cytokines have been directly correlated with GvHD pathogenesis [42]–[44], early post-transplant administration of rflagellin in allo-BMT could provide clinical benefit in allo-BMT recipients by enhancing NK cell activity [45] without increasing the risk of GvHD.…”

Activation of Innate Immunity in Graft-versus-Host Disease: Implications for Novel Targets?

“…However, the role of TLRs in GvHD is complicated by the fact that also tolerogenic innate immune cells may depend on TLR-based activation, such as for example myeloid-derived suppressor cells which are potent regulators of GvHD [5]. Consistent with a regulatory role of certain TLRs, pretreatment of mice with the TLR5 ligand flagellin resulted in reduced GvHD severity [6]. Administration of an agonist to TLR-7/8 induced indoleamine 2,3-dioxygenase (IDO) and reduced GvHD-related injury in the colon and ameliorated lethality [7].…”